Daniel:Notebook/HiResChrPaint/2014-6-19

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Probe Design (Started 06/17/2014)[edit]

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Probe Mining[edit]

The OligoArray script might actually need the blast database broken up into 1kb chunks for some reason. So, let's try that.

  1. Create BLAST database from fragmented genome
    1. formatdb -i in_hg38.fas -p T -o F
[formatdb] WARNING: Cannot add sequence number 1953021 (lcl|range=hg38:1953020000-1953020999) because it has zero-length.
[formatdb] FATAL ERROR: Fatal error when adding sequence to BLAST database.

So that's a fatal error.

The instructions call for concatenating all the fasta files into one. I did this step using cat, but they suggest using their java script, so I'll try that.

  1. Concatenate fasta files
    1. java ConcatenateFiles hg38_cat.fas *.fa
  2. Fragment genome into pieces
    1. python ../bin/input_blocks.py
    2. Please enter the filename, "chr2R_sorted_oligo.txt" or "chr2R_intersect.bed" etc.: hg38_cat.fas
    3. Please enter the chromosome or assembly, "chr1 or ch2R, etc.: hg38
Killed

Couldn't have said it better myself.

Trying again using the Wu lab (Oligopaints) version of the fasta file. This is an older version (hg19/gr37, not gr38), but it'll have to do.

  1. Format BLAST database
    1. formatdb -i hg19.fas -p F -o T
  2. Run OligoArray2
    1. java -Xmx8192m -jar /home/djacobse/blastdb_hg38/OligoArray2_1/OligoArray2.jar -i in_caskgene.fas -d hg19.fas -o oligo_caskgene.txt -r rejected_oligo.fas -R caskgene.log -n 30 -l 60 -L 60 -D 1000 -t 55 -T 65 -s 70 -x 70 -p 35 -P 80 -m "CCCC;GGGG;TTTTT;AAAAA" -g 2
Start range=chrX:1157000-1157999
Running Blast (-e 0.0)... For sequence range=chrX:1157000-1157999, the following exception was caugth:
java.io.IOException: Cannot run program "blastall": java.io.IOException: error=12, Cannot allocate memory

New problem, sort of. Looks like it can run OligoArray for a little while (gets through about 5000 bp or so), then says it can't allocate memory. Not sure what that's all about.

I may be allocating more memory than is available for allocation with this command, so I'll try again without the memory allocation flag. I may also need to try again using a different (smaller) memory allocation flag.

  1. Run OligoArray2
    1. java -jar /home/djacobse/blastdb_hg38/OligoArray2_1/OligoArray2.jar -i in_caskgene.fas -d hg19.fas -o oligo_caskgene.txt -r rejected_oligo.fas -R caskgene.log -n 30 -l 60 -L 60 -D 1000 -t 55 -T 65 -s 70 -x 70 -p 35 -P 80 -m "CCCC;GGGG;TTTTT;AAAAA" -g 2